Comparison of biochemical and biological effects of four nitrosoureas with differing carbamoylating activities.

Four chloroethylnitrosoureas with differing degrees of carbamoylating activity were compared for their effects on incorporation of radioactive precursors into macromolecules. The comparisons were made with concentrations that, for each drug, produced a defined biological effect, either an 0.5-log or a 2-log reduction in cloning efficiency from a 1-hr drug exposure. Dose-dependent inhibition of incorporation of labeled precursors into nucleic acids and proteins was observed during the 1-hr drug exposure with either of the two strongly carbamoylating nitrosoureas, 1-3-bis(2-chloroethyl)-u-nitrosourea and 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea; the most extensive inhibition involved incorporation into DNA. No inhibitions were observed during exposure to a weakly carbamoylating nitrosourea (chlorozotocin) or during exposure to 1-(2-chloroethyl)-1-nitrosourea, a compound the carbamoylating activity of which is not agreed upon. By 4 hr after removal of 1-3-bis(2-chloroethyl)-1-nitrosourea or 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea from the extracellular medium, the cells had partially recovered from the earlier inhibition of radioactive thymidine incorporation. This recovery was, however, dependent upon an undefined factor present in serum. The inhibitions that were observed during the 1-hr drug exposure are clearly not essential to the cytotoxic effect of chloroethylnitrosoureas since no inhibitions occurred with two of the four compounds studied.

DNA single-strand breaks during repair of UV damage in human fibroblasts and abnormalities of repair in xeroderma pigmentosum.

The method of DNA alkaline elution was applied to a study of the formation and resealing of DNA single-strand breaks after irradiation of human fibroblasts with ultraviolet light (UV). The general features of the results were consistent with current concepts of DNA excision repair, in that breaks appeared rapidly after UV, and resealed slowly in normal fibroblasts, whereas breaks did not appear in those cells of patients with xeroderma pigmentosum (XP) that are known to have defects in DNA repair synthesis. The appearance of breaks required a short post-UV incubation, consistent with the expected action of an endonuclease. Cells of the variant form of XP characterized by normal DNA repair synthesis exhibited normal production of breaks after UV, but were slower than normal cells in resealing these breaks. This difference was enhanced by caffeine. A model is proposed to relate this finding with a previously described defect in post-replication repair in these XP variant cells. DNA crosslinking appears to cause an underestimate in the measurement of DNA breakage after UV.

Studies of paroxysmal nocturnal hemoglobinuria erythrocytes: increased lysis and lipid peroxide formation by hydrogen peroxide.

When paroxysmal nocturnal hemoglobinuria (PNH) erythrocytes were exposed to H(2)O(2) they lysed excessively and formed greater than normal quantities of lipid peroxides when compared to red cells of normal subjects and patients with most types of hematologic disease. It was also shown that lytic sensitivity to acidified serum was related to the enhanced lytic sensitivity to H(2)O(2). If the lipid of PNH cells was first extracted then exposed to ultraviolet radiation more lipid peroxides were formed than in extracts of normal red blood cells. The possible explanations for these findings and their relationship to the PNH hemolytic mechanism are discussed.